Neurons communicate via electrical impulses, called action potentials, that last for less than one thousandth of a second and travel in fibers, called axons, approximately one micrometer in diameter. The small size of axons has prevented direct characterization of action potentials with traditional recording methods. I will utilize recent advancements in single cell voltage-sensitive dye (VSD) imaging to measure changes in membrane electrical potential throughout the neuron with high resolution in time and space. To obtain a better understanding of how neurons process and communicate information, I will use this powerful new technique to image initiation and propagation of single action potentials throughout the axonal arbor of cerebellar Purkinje neurons. In Specific Aim 1, I will directly measure the site of action potential initiation. This information is fundamental to our understanding of how neurons respond to their inputs. In Specific Aim 2, I will investigate whether action potentials travel faithfully throughout the axonal arbor. Earlier theory and experiments have shown that the transmission of information between connected neurons is stochastic, and failures in action potential transmission through highly branched axons have been hypothesized as a mechanism. My direct observations of action potential transmission with VSD imaging will show where and how often these failures occur. In Specific Aim 3, I will investigate how the recent history of input to a neuron influences action potential shape and transmission. Previous experiments have shown that recent inputs to a neuron can influence how it communicates with neighboring cells. I will use VSD imaging to observe how recent history of inputs influences action potential shape and transmission fidelity throughout the axonal arbor. These experiments will increase our understanding of the fundamental role played by axons in neuronal communication and should provide insights into the development and progression of neurological syndromes that arise from abnormal changes in axonal function. PUBLIC HEALTH RELEVANCE: The proper function of axons, the primary output structures of neurons, is a critical component of a healthy, functional brain and nervous system. Our study will use recently advanced technologies to elucidate important basic properties of normal axonal physiology. Our data will provide new insights into how neurological symptoms and syndromes, such as multiple sclerosis and peripheral neuropathies, may arise from abnormal changes in axonal function.